Synthesis and secretion of the extracellular domain of the β1 subunit isoform of NA⁺/K⁺-ATPase into the culture medium of CHO cells

Abstract

The membrane topology of the β subunit of Na⁺/K⁺-ATPase suggests that the protein has one single membrane spanning segment about 35 amino-acids behind the N-terminus, followed by a huge extracellular domain. Therefore, a recombinant α1 subunit protein lacking the intracellular N-terminal amino-acids as well as the computer predicted transmembrane domain should be directed to the secretory pathway if provided with the appropriate signal peptide sequences at the new N-terminal end. We have chosen this approach for two reasons. First, secretion into the culture medium would clearly demonstrate that the β subunit of the Na⁺/K⁺- ATPase β subunit is anchored by one single transmembrane segment. Second, since structural analysis of membrane anchored proteins is still a difficult task to solve, the secreted extracellular domain could be a suitable target for such analysis as has been shown for example with the extracellular domain of CD4 (1).